We propose to undertake a detailed genetic analysis of Sindbis virus, a group A arbovirus. These viruses consist of an icosahedral nucleocapsid containing the viral nucleic acid (a single-stranded RNA with a molecular weight of 4.2 million daltons) surrounded by a lipoprotein envelope or membrane. The viral genome probably contains 10 to 12 cistrons, of which five are currently known. We plan to isolate a large number of temperature- sensitive mutants after chemical mutagenesis with a number of agents. Using these conditional-lethal mutants, as well as those isolated previously by Dr. B.W. Burge, we hope to enumerate the total number of cistrons by complementation tests and to determine the function of each gene. A wide variety of biochemical and biophysical assays will be used to study gene functions, including examination of the RNA species made during infection, detailed analysis of the mutant proteins produced (by acrylamide gel electrophoresis), chemical and physical studies of the mutant virions and cores, and temperature shift experiments. We are especially interested in mutants which synthesize altered membrane components. These will be studied immunologically to locate altered antigenic determinants, by determination of the extent of integration of the altered envelope protein into the cell membrane, examination of the carbohydrate chains of this glycoprotein, and studying the association of cores with the surface membrane.